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Toxins 2017, 9(12), 382; https://doi.org/10.3390/toxins9120382

Multipurpose HTS Coagulation Analysis: Assay Development and Assessment of Coagulopathic Snake Venoms

1
Division of BioAnalytical Chemistry, Department of Chemistry and Pharmaceutical Sciences, Faculty of Sciences, Vrije Universiteit Amsterdam, De Boelelaan 1085, 1081 HV Amsterdam, The Netherlands
2
Alistair Reid Venom Research Unit, Parasitology Department, Liverpool School of Tropical Medicine, Liverpool L3 5QA, UK
3
Research Centre for Drugs and Diagnostics, Liverpool School of Tropical Medicine, Liverpool L3 5QA, UK
*
Author to whom correspondence should be addressed.
Academic Editor: Frank S. Markland
Received: 20 October 2017 / Revised: 16 November 2017 / Accepted: 21 November 2017 / Published: 25 November 2017
(This article belongs to the Section Animal Venoms)
View Full-Text   |   Download PDF [4045 KB, uploaded 25 November 2017]   |  

Abstract

Coagulation assays currently employed are often low throughput, require specialized equipment and/or require large blood/plasma samples. This study describes the development, optimization and early application of a generic low-volume and high-throughput screening (HTS) assay for coagulation activity. The assay is a time-course spectrophotometric measurement which kinetically measures the clotting profile of bovine or human plasma incubated with Ca2+ and a test compound. The HTS assay can be a valuable new tool for coagulation diagnostics in hospitals, for research in coagulation disorders, for drug discovery and for venom research. A major effect following envenomation by many venomous snakes is perturbation of blood coagulation caused by haemotoxic compounds present in the venom. These compounds, such as anticoagulants, are potential leads in drug discovery for cardiovascular diseases. The assay was implemented in an integrated analytical approach consisting of reversed-phase liquid chromatography (LC) for separation of crude venom components in combination with parallel post-column coagulation screening and mass spectrometry (MS). The approach was applied for the rapid assessment and identification of profiles of haemotoxic compounds in snake venoms. Procoagulant and anticoagulant activities were correlated with accurate masses from the parallel MS measurements, facilitating the detection of peptides showing strong anticoagulant activity. View Full-Text
Keywords: snake venoms; blood coagulation; coagulation bioassay; high-throughput screening; mass spectrometry; liquid chromatography snake venoms; blood coagulation; coagulation bioassay; high-throughput screening; mass spectrometry; liquid chromatography
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Still, K.B.M.; Nandlal, R.S.S.; Slagboom, J.; Somsen, G.W.; Casewell, N.R.; Kool, J. Multipurpose HTS Coagulation Analysis: Assay Development and Assessment of Coagulopathic Snake Venoms. Toxins 2017, 9, 382.

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